Coordinated Regulation of Myelination by Growth Factor and Amino-acid Signaling Pathways / 神经科学通报·英文版

Myelin-forming oligodendrocytes in the central nervous system (CNS) and Schwann cells in the peripheral nervous system (PNS) are essential for structural and functional homeostasis of nervous tissue. Albeit with certain similarities, the regulation of CNS and PNS myelination is executed differently. Recent advances highlight the coordinated regulation of oligodendrocyte myelination by amino-acid sensing and growth factor signaling pathways. In this review, we discuss novel insights into the understanding of differential regulation of oligodendrocyte and Schwann cell biology in CNS and PNS myelination, with particular focus on the roles of growth factor-stimulated RHEB-mTORC1 and GATOR2-mediated amino-acid sensing/signaling pathways. We also discuss recent progress on the metabolic regulation of oligodendrocytes and Schwann cells and the impact of their dysfunction on neuronal function and disease.

Effect of Rheb1 in the Development of Mouse Megakaryocyte-Erythroid Progenitor Cells / 中国实验血液学杂志

OBJECTIVE@#To investigate the effect of Rheb1 in the development of mouse megakaryocyte-erythroid progenitor cells and its related mechanism.@*METHODS@#Rheb1 was specifically knocked-out in the hematopoietic system of Vav1-Cre;Rheb1fl/fl mice(Rheb1Δ/Δ mice). Flow cytometry was used to detect the percentage of red blood cells in peripheral blood and erythroid cells in bone marrow in Vav1-Cre;Rheb1fl/fl mice and control mice. The CFC assay was used to detect the differentiation ability of Rheb1 KO megakaryocyte-erythroid progenitor cells and control cells. Real-time fluorescence quantification PCR was used to detect the relative expression of PU.1,GATA-1,GATA-2,CEBPα and CEBPβ of Rheb1 KO megakaryocyte-erythroid progenitor cells and control cells. Rapamycin was added to the culture medium, and it was used to detect the changes in cloning ability of megakaryocyte-erythroid progenitor cells from wild-type mice in vitro.@*RESULTS@#After Rheb1 was knocked out, the development and stress response ability of megakaryocyte-erythroid progenitor cells in mice were weaken and the differentiation ability of megakaryocyte-erythroid progenitor cells in vitro was weaken. Moreover, the expression of GATA-1 of megakaryocyte-erythroid progenitor cells was decreased. Further, rapamycin could inhibit the differentiative capacity of megakaryocyte-erythroid progenitor cells in vitro.@*CONCLUSION@#Rheb1 can regulate the development of megakaryocyte-erythroid progenitor cells probably through the mTOR signaling pathway in mice.

Farnesylation-defective Rheb Increases Axonal Length Independently of mTORC1 Activity in Embryonic Primary Neurons

Rheb (Ras homolog enriched in the brain) is a small GTPase protein that plays an important role in cell signaling for development of the neocortex through modulation of mTORC1 (mammalian-target-of-rapamycin-complex-1) activity. mTORC1 is known to control various biological processes including axonal growth in forming complexes at the lysosomal membrane compartment. As such, anchoring of Rheb on the lysosomal membrane via the farnesylation of Rheb at its cysteine residue (C180) is required for its promotion of mTOR activity. To test the significance of Rheb farnesylation, we overexpressed a farnesylation mutant form of Rheb, Rheb C180S, in primary rat hippocampal neurons and also in mouse embryonic neurons using in utero electroporation. Interestingly, we found that Rheb C180S maintained promotional effect of axonal elongation similar to the wild-type Rheb in both test systems. On the other hand, Rheb C180S failed to exhibit the multiple axon-promoting effect which is found in wild-type Rheb. The levels of phospho-4EBP1, a downstream target of mTORC1, were surprisingly increased in Rheb C180S transfected neurons, despite the levels of phosphorylated mTOR being significantly decreased compared to control vector transfectants. A specific mTORC1 inhibitor, rapamycin, also could not completely abolish axon elongation characteristics of Rheb C180S in transfected cells. Our data suggests that Rheb in a non-membrane compartment can promote the axonal elongation via phosphorylation of 4EBP1 and through an mTORC1-independent pathway.

Analysis of Rheb in the cellular slime mold Dictyostelium discoideum: Cellular localization, spatial expression and overexpression.

Dictyostelium discoideum encodes a single Rheb protein showing sequence similarity to human homologues of Rheb. The DdRheb protein shares 52% identity and 100% similarity with the human Rheb1 protein. Fluorescence of Rheb yellow fluorescent protein fusion was detected in the D. discoideum cytoplasm. Reverse transcription-polymerase chain reaction and whole-mount in situ hybridization analyses showed that rheb is expressed at all stages of development and in prestalk cells in the multicellular structures developed. When the expression of rheb as a fusion with lacZ was driven under its own promoter, the β-galactosidase activity was seen in the prestalk cells. D. discoideum overexpressing Rheb shows an increase in the size of the cell. Treatment of the overexpressing Rheb cells with rapamycin confirms its involvement in the TOR signalling pathway.

Role of a regulating protein in the upper stream of mTOR signaling pathway Rheb in adipocyte differentiation / 第二军医大学学报

Objective To observe the effect of Rheb, a regulating protein in the upper stream of mTOR signaling pathway, on adipocyte differentiation. Methods (1) The recombinant plasmid pCAG-Insulator-Rheb was constructed and injected into the embryo of B6 mouse to produce transgenic mouse over-expressing Rheb. (2) The mouse embryonic fibroblast cells (MEFs) were collected from Rheb transgenic mice on pregnancy day 13. 5, and they were induced to differentiate into adipocytes after identified by PCR. (3) On differentiation day 12, the MEFs were subjected to Oil Red O staining and the contents of triglyceride in the MEFs were determined; moreover, the expression of special transcription factor PPARy and C/ EBPa was examined by real-time PCR, and all of the above parameters were used to assess the role of Rheb in the differentiation of adipocytes. Results We successfully constructed the transgenic mouse model over-expressing Rheb. We found that mouse over-expressing Rheb promoted adipogenesis in MEFs and increased the content of triglyceride in MEFs; moreover, the overexpression also greatly changed the results of Oil Red O staining and greatly promoted the expression of adipocyte specific transcript factor PPARy and C/EBPα. Conclusion Over-expression of Rheb can promote the differentiation of MEFs into adipocytes.

Expressional Difference of RHEB, HDAC1, and WEE1 Proteins in the Stromal Tumors of the Breast and Their Significance in Tumorigenesis

BACKGROUND: Fibroadenoma (FA) and phyllodes tumor (PT) are stromal tumors of breast and are histologically similar. There are no established differences in tumorigenesis and oncogene expression among them. Ras homolog enriched in brain (RHEB) plays an important role in cell growth and cell-cycle control, histone deacetylase 1 (HDAC1) is an important factor in breast tumor progression and prognosis, and WEE1 homolog (WEE1) functions as a tumor suppressor. No studies on the expressional differences of these proteins in FA and PT have been reported to date. METHODS: The expression of these proteins in FA, PT, and normal breast was compared. We used 102 cases of FA and 25 cases of benign PT. RESULTS: In epithelial cells, the expression of RHEB, HDAC1, and WEE1 was lowest in PT, higher in FA, and most enhanced in normal breast. In addition, the expression of RHEB and HDAC1 was higher in the stromal cells of PT than in FA and normal breast. CONCLUSIONS: Both epithelial and stromal cells of FA and PT express these proteins, which indicates that epithelial cells play an important role in the development of stromal tumors. In addition, the expressional differences of these proteins may be associated with the tumorigenesis of breast stromal tumors.